Application of Plant Extracts for the Prevention of Dental Erosion: An in situ/in vitro Study.

OBJECTIVES: Antiadherent and antibacterial effects of certain plant extracts have been proven to be beneficial in preventive dentistry. In the present in situ/in vitro crossover study, the impact of plant extracts rich in polyphenols on the erosion-protective properties of the in situ pellicle was evaluated. METHODS: Individual splints were prepared for 12 subjects for intraoral exposure of bovine enamel specimens. Following formation of a 1-min pellicle, watery plant extracts (leaves of the wild form of Ribes nigrum, the wild form of Origanum as well as a combination of both) were administered for 10 min in situ. Alternatively, a mouth rinse with fluorides (Elmex Kariesschutz) was performed for 1 min. After further oral exposure for 19/28 min, respectively, slabs were removed and incubated with HCl in vitro over 120 s (pH 2, 2.3, 3). The resulting calcium and phosphate release was quantified photometrically. Slabs with and without a 30-min in situ pellicle served as controls. The modification of pellicle ultrastructure was evaluated by transmission electron microscopy (TEM). RESULTS: Plant extracts modulated the erosion-protective properties of the native in situ pellicle in all test groups in a pH-dependent manner. The combination of R. nigrum leaves and Origanum enhanced the protective properties of the pellicle at all pH values; the administration of this preparation was comparable, yet superior, to the effect of the fluoridated mouth rinse. TEM images indicated that rinsing with R. nigrum leaves/Origanum yielded a distinctly thicker and more electron-dense pellicle. CONCLUSION: The combination of certain plant extracts offers a novel approach to the complementary prevention of dental erosion.

Colour improvement and stability of white spot lesions following infiltration, micro-abrasion, or fluoride treatments in vitro.

SUMMARY BACKGROUND/OBJECTIVES: White spot lesions (WSLs) are unwelcome side effects of fixed appliances that compromise the treatment outcome. Recently, infiltration of WSLs has been introduced as a viable treatment alternative. The objective was to evaluate the colour improvement of WSLs and their stability against discolouration following infiltration, fluoride, or micro-abrasion treatments in vitro. MATERIALS/METHODS: Artificial WSLs were created in bovine enamel (N = 96) using acidic buffer solution (pH 5, 10 days) and were randomly allocated to four groups. Specimens were treated with infiltration (Icon, DMG), fluoride (Elmex Caries Protection, GABA), and micro-abrasion (Opalustre, Ultradent) or remained untreated (control). Groups were discoloured for 24 hours in tea or tea + citric acid. Colour components and visible colour change (L*, a*, b*, ΔE) were measured spectrophotometrically on following time points: baseline, after WSL formation, after treatment, and during discolouration (8, 16, and 24 hours). Data were analysed using Kruskal-Wallis and Mann-Whitney tests. RESULTS: WSL formation increased (L*) in all groups. Only infiltration reduced this effect to baseline. Highest ΔE improvement was obtained by infiltration and micro-abrasion followed by fluoride. This improvement was stable only for infiltration during discolouration. L*, a*, and b* changed significantly during discolouration in all groups except infiltration. Within the same treatment group, discolouration solutions did not differ significantly. LIMITATIONS: In vitro testing cannot replicate the actual mode of colour improvement or stability but can be used for ranking materials and techniques. CONCLUSIONS/IMPLICATIONS: Infiltration and micro-abrasion treatments were capable of diminishing the whitish appearance of WSLs. Only infiltrated WSLs were stable following discolouration challenge.

Effect of conventional mouthrinses on initial bioadhesion to enamel and dentin in situ.

AIM: The study aimed to investigate the effect of a customary fluoride solution, containing sodium fluoride and amine fluoride, on initial biofilm formation on enamel and dentin in situ compared directly to chlorhexidine. METHODS: Bovine enamel and dentin specimens were mounted on maxillary splints carried by 9 subjects. After 1 min of pellicle formation, rinses with tap water (control), chlorhexidine (meridol med CHX 0.2%, GABA) and a fluoride mouthrinse (elmex, GABA) were performed for 1 min. Subsequently, the slabs were carried for another 8 h. The adherent bacteria were determined by DAPI staining, live-dead staining and determination of colony-forming units after desorption; glucan formation was visualized with concanavalin A. Additionally, energy-dispersive X-ray spectroscopy (EDX) analysis of the in situ biofilm layers was conducted, and contact angle measurements were performed. Statistical evaluation was performed by means of the Kruskal-Wallis test followed by the Mann-Whitney U test (p < 0.05). RESULTS: In the control group, significantly higher amounts of adherent bacteria were detected on dentin (4.8 × 10(6) ± 5.4 × 10(6) bacteria/cm(2)) than on enamel (1.2 × 10(6) ± 1.5 × 10(6) bacteria/cm(2), DAPI). Chlorhexidine significantly reduced the amount of adherent bacteria (dentin: 2.8 × 10(5) ± 3.4 × 10(5) bacteria/cm(2); enamel: 4.2 × 10(5) ± 8.7 × 10(5) bacteria/cm(2)). Rinses with the fluoride solution also significantly reduced bacterial adherence to dentin (8.1 × 10(5) ± 1.5 × 10(6) bacteria/cm(2)). Fluoride could not be detected by EDX analysis of the biofilms. Fluoride mouthrinsing did not influence the wettability of the pellicle-covered enamel surface. CONCLUSION: In addition to the reduction of demineralization and antibacterial effects, fluorides inhibit initial biofilm formation on dental hard tissues considerably, especially on dentin.

Clinical effects of an essential oil solution used as a coolant during ultrasonic root debridement.

AIM: The use of chlorhexidine and povidone iodine solutions applied as a coolant during ultrasonic root debridement for the treatment of chronic periodontitis has been described. Hitherto, this application has not yet been extensively investigated for essential oil solutions. The goal was to clinically explore this and to compare to water irrigation. MATERIALS AND METHODS: Thirty-five chronic periodontitis patients participated in a single-blind randomized controlled clinical study. Patients were randomly allocated to the control group (n=18) or test group (n=17) receiving oral hygiene instructions and ultrasonic root debridement using water as a coolant, respectively, a pure essential oil solution. Oral hygiene was reinforced if necessary at each occasion, and clinical parameters were collected at baseline and after 1 and 3 months. RESULTS: Significant pocket reduction (control, 1.02 mm; test, 0.89 mm) and clinical attachment gain (control and test, 0.48 mm) were shown in both groups. However, there were no significant differences between the groups at any point in time for any of the parameters. CONCLUSION: Essential oil solutions do not offer a clinical benefit over water when used as a coolant during ultrasonic root debridement for the treatment of chronic periodontitis.

[Effects of ionotropic glutamate receptor channel blockers on the development of audiogenic seizures in Krushinski-Molodkina rats].

The action of noncompetitive blockers of glutamate receptors has been investigated on Krushinski-Molodkina rats genetically-prone to audiogenic seizures. The selective blockers of NMDA receptor channels, memantine and IEM-1921, and their dicationic homologues, IEM-1925 and IEM-1754, capable of blocking in varying degrees both NMDA and Ca-permeable AMPA receptor channels, were studied. The drugs were injected intramuscularly to rats with the different time intervals (30 min, 1, 2 or 3 hours) before sound signal. The effects of the drugs on latent period of initial locomotor activity provoked by audio stimulation (8 kHz sine-wave tone, 90 dB volume), the appearance of clonic convulsions of different intensities, and, finally, tonic convulsions with limb and tail extension were evaluated. Within 30 min after injection IEM-1921 at a dose of 5 mg/kg, 33% of rats manifested a complete absence of convulsive reactions to sound, and in 59% of rats audiogenic seizures occured only in the form of motor excitation without a generalized clonic-tonic convulsions. Memantine at a dose of 5 mg/kg did not cause a complete blockade of seizures, but after 1 h of injection in 50% of the rats and after 2 h in 70% of rats a weakening of the audiogenic seizures to the level of motor excitation only was observed. After 3 hrs after administration of blockers its anticonvulsive action weakened significantly (p < 0.01). Dicationic blockers that block both NMDA and AMPA/kainate receptors, IEM-1925 (in doses of 0.001-20.0 mg/kg) and IEM-1754 (0.025-50.0 mg/kg), did not affect audiogenic clonic-tonic convulsive reactions. The involvement of activation of NMDA and calcium permeable AMPA/kainate receptors in the pathogenesis of audiogenic seizures is discussed.

Prevention of toothbrushing abrasion of acid-softened enamel by CO(2) laser irradiation.

OBJECTIVE: The aim of the present study was to evaluate the effect of CO(2) laser irradiation (10.6µm) at 0.3J/cm(2) (0.5µs; 226Hz) on the resistance of softened enamel to toothbrushing abrasion, in vitro. METHODS: Sixty human enamel samples were obtained, polished with silicon carbide papers and randomly divided into five groups (n=12), receiving 5 different surface treatments: laser irradiation (L), fluoride (AmF/NaF gel) application (F), laser prior to fluoride (LF), fluoride prior to laser (FL), non-treated control (C). After surface treatment they were submitted to a 25-day erosive-abrasive cycle in 100ml sprite light (90s) and brushed twice daily with an electric toothbrush. Between the demineralization periods samples were immersed in supersaturated mineral solution. At the end of the experiments enamel surface loss was determined using a contact profilometer and morphological analysis was performed using scanning electron microscopy (SEM). For SEM analysis of demineralization pattern, cross-sectional cuts of cycled samples were prepared. The data were statistically analysed by one-way ANOVA model with subsequent pairwise comparison of treatments. RESULTS: Abrasive surface loss was significantly lower in all laser groups compared to both control and fluoride groups (p<0.0001 in all cases). Amongst the laser groups no significant difference was observed. Softened enamel layer underneath lesions was less pronounced in laser-irradiated samples. CONCLUSION: Irradiation of dental enamel with a CO(2) laser at 0.3J/cm(2) (5µs, 226Hz) either alone or in combination with amine fluoride gel significantly decreases toothbrushing abrasion of softened-enamel, in vitro.

Rehardening of acid-softened enamel and prevention of enamel softening through CO2 laser irradiation.

OBJECTIVES: The aims of the present study were to investigate whether irradiation with a CO(2) laser could prevent surface softening (i) in sound and (ii) in already softened enamel in vitro. METHODS: 130 human enamel samples were obtained and polished with silicon carbide papers. They were divided into 10 groups (n = 13) receiving 5 different surface treatments: laser irradiation (L), fluoride (AmF/NaF gel) application (F), laser prior to fluoride (LF), fluoride prior to laser (FL), non-treated control (C); and submitted to 2 different procedures: half of the groups was acid-softened before surface treatment and the other half after. Immersion in 1% citric acid was the acid challenge. Surface microhardness (SMH) was measured at baseline, after softening and after treatment. Additionally, fluoride uptake in the enamel was quantified. The data were statistically analysed by two-way repeated measurements ANOVA and post hoc comparisons at 5% significance level. RESULTS: When softening was performed either before or after laser treatment, the L group presented at the end of the experiments SMH means that were not significantly different from baseline (p = 0.8432, p = 0.4620). Treatment after softening resulted for all laser groups in statistically significant increase in SMH means as compared to values after softening (p < 0.0001). Enamel fluoride uptake was significantly higher for combined laser-fluoride treatment than in control (p<0.0001). CONCLUSION: Irradiation of dental enamel with a CO(2) laser at 0.3J/cm(2) (5 µs, 226 Hz) not only significantly decreased erosive mineral loss (97%) but also rehardened previously softened enamel in vitro.

Effect of various rinsing protocols after use of amine fluoride/stannous fluoride toothpaste on the bacterial composition of dental plaque.

This clinical study evaluated the effect of different oral hygiene protocols on the bacterial composition of dental plaque. After a 2-week period of using fluoride-free toothpaste, 30 participants followed three 1-week experimental protocols, each followed by 2-week fluoride-free washout periods in a randomized crossover examiner-blind controlled trial. The 1-week experimental protocols comprised the use of AmF/SnF(2) toothpaste twice daily, after which participants either (1) rinsed with tap water, (2) did not rinse but only spat out the toothpaste, or (3) rinsed with an AmF/SnF(2) mouthwash. In the fluoride-free washout periods, the participants brushed their teeth with fluoride-free toothpaste without further instructions. Six hours after the last brushing (+/- rinsing) of each period, buccal plaque samples in the upper molar region were taken. The microbiota composition of the plaque samples was analyzed by checkerboard DNA:DNA hybridization. A statistically significant reduction was found in the total amount of DNA of the 39 major plaque species measured, and in the proportions of some acid-producing bacterial strains after the period having used the AmF/SnF(2) toothpaste + AmF/SnF(2) mouthrinsing. The results indicate that using the AmF/SnF(2) toothpaste and rinse combination could result in plaque of lower cariogenicity.

Recent advances in the preclinical evaluation of the topical antibacterial agent REP8839.

REP8839 is a synthetic fluorovinylthiophene-containing diaryldiamine that inhibits bacterial methionyl tRNA synthetase (MetRS) and is a new chemical entity that represents a novel pharmacological class. The compound has potent in vitro antibacterial activity against many clinically important Gram-positive bacteria including the major skin pathogens Staphylococcus aureus and Streptococcus pyogenes. In light of the emergence of methicillin-resistant S. aureus in the community and increasing resistance to mupirocin, REP8839 is being evaluated as a topical agent for the treatment of superficial skin infections. REP8839 was active against resistant phenotypes of S. aureus and can be formulated at high concentrations to minimize the development of resistance. A formulation of REP8839 has demonstrated efficacy in a porcine partial thickness wound infection model against mupirocin-resistant S. aureus.

Impact of fluoride, milk and water rinsing on surface rehardening of acid softened enamel. An in situ study.

PURPOSE: To evaluate the impact of fluoride, milk and water rinsing on surface rehardening of acid softened enamel in situ. METHODS: Ten subjects performed six tests of4 hours each. In each test, three softened enamel samples were attached to intraoral appliances. For softening, samples were immersed extraorally in an acidic beverage for 120 seconds. Subsequently, specimens were worn intraorally for 5 minutes (Tests 1-3) or 30 minutes (Tests 4-6). Thereafter, the volunteers rinsed with a 250 ppm SnF2/Olaflur solution (Tests 1 and 4), milk (Tests 2 and 5) or non-carbonated mineral water (Tests 3 and 6) for 60 seconds. At each test, one sample was covered with tape during intraoral rinsing and thus, served as control. After rinsing, both test and the control samples were exposed to the oral cavity for up to 4 hours after demineralization. Surface microhardness (SMH) of the specimens was assessed at baseline, immediately after softening and 4 hours after softening. For each subject, the secretion rate of resting and stimulated saliva, buffering capacity and pH-value as well as calcium and phosphate concentration of saliva were measured. Statistical analysis was performed by ANCOVA followed by stratified analyses with Bonferroni correction. RESULTS: Baseline Knoop Hardness (mean +/- S.D.) amounted to 403.1 +/- 39.4. Immediately after softening, mean SMH was reduced to 214.4 +/- 24.1 KHN. Rinsing with 250 ppm fluoride, milk or water after 5 minutes or 30 minutes intraoral exposure of softened samples had a significant effect on rehardening. The increase of SMH (DeltaKHN) was highest after rinsing with fluoride (5 minutes: 95.0 +/- 18.3; 30 minutes: 94.2 +/- 24.3) followed by milk (5 minutes: 77.1 +/- 14.1; 30 minutes: 80.3 +/- 18.7) and water (5 minutes: 49.0 +/- 9.9; 30 minutes: 47.0 +/- 14.1), but did not achieve baseline values. It is concluded that a single rinse with a 250 ppm SnF2/Olaflur solution, milk or water increases rehardening of previously acid softened enamel.

Inhibitory effects of 1,3-diaminopropane, an ornithine decarboxylase inhibitor, on rat two-stage urinary bladder carcinogenesis initiated by N-butyl-N-(4-hydroxybutyl)nitrosamine.

Overexpression of ornithine decarboxylase (ODC) has been shown to be characteristic of tumor development and progression in humans and experimental animals. Therefore, we have examined the effects of 1, 3-diaminopropane dihydrochloride (DAP), a potent inhibitor of ODC, on rat two-stage urinary bladder carcinogenesis initiated with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN). In experiment 1 (36 weeks), 6-week-old F344 male rats were administered 0.05% BBN in drinking water for 4 weeks and then divided into four groups. Animals of groups 1 and 2 received basal diet and drinking water supplemented with or without DAP (2 g/l). Groups 3 and 4 were given diet containing 5% sodium L-ascorbate (NaAsA), a typical urinary bladder tumor promoter, and drinking water with or without DAP. Administration of DAP to group 1 significantly reduced tumor size, multiplicity and incidence, particularly of papillomas, when compared with group 2 values. DAP together with NaAsA (group 3) also decreased tumor size relative to the group 4 case. To determine the effects of DAP on the early stages of bladder carcinogenesis and its mechanisms, a similar protocol was conducted (experiment 2) with death after 20 weeks. DAP treatment caused complete inhibition (0% incidence) of papillary and/or nodular hyperplasia in group 1 but was without influence in group 3, as compared with the respective controls. Moreover, the ODC activity, bromodeoxyuridine labeling indices and mRNA expression levels of cyclin D1 in the urinary bladder mucosa, determined by northern blotting, were markedly lower in group 1 than in group 2, but values were comparable for both groups administered NaAsA. Assessment of mRNA expression levels of the angiogenic vascular endothelial growth factor suggested no involvement in the inhibitory effects of DAP on urinary bladder carcinogenesis. The results indicate that inhibition of ODC could reduce urinary bladder carcinogenesis in rats, particularly in the early stages, through antiproliferative mechanisms.

Effects of glutathione or polyamine depletion on in vivo thermosensitization.

Investigations with the melphalan-sensitive and -resistant human rhabdomyosarcoma xenografts TE-671 and TE-671 MR were performed to examine the effect of glutathione and polyamine modulation on thermosensitivity. Regimens of intraperitoneally injected and orally administered buthionine sulfoximine were utilized to achieve glutathione depletion to 8.7% and 13% of control levels in TE-671 and TE-671 MR, respectively. Animals treated with L-buthionine-S,R-sulfoximine and 42 degrees C or 43 degrees C hyperthermia for 70 min showed no detectable growth delays beyond those observed for hyperthermia alone. Hyperthermia at 42 degrees C of disaggregated TE-671 and TE-671 MR xenografts following growth in short-term culture was performed following preincubation with buthionine sulfoximine or 0.9% saline. Buthionine sulfoximine-mediated glutathione depletion produced a significant increase in hyperthermia-induced cytotoxicity only with TE-671 MR at 43 degrees C. Polyamine depletion was achieved with a 7-day orally administered course of MDL 72.175DA [(2R,5R)-6-heptyne,5-diamine dihydrochloride], an irreversible inhibitor of ornithine decarboxylase. Although this treatment caused significant depletion of intracellular putrescine and spermidine levels, spermine levels remained relatively unaffected. No significant growth delays were observed in either xenograft line for animals treated with MDL 72.175DA or MDL 72.175DA plus hyperthermia as compared with untreated controls. These results contrast with previous work performed in vitro showing synergism between glutathione or polyamine depletion and hyperthermia, and indicate that further studies are needed.

The antitumoral action of polyamine linked cyclophosphazenes on human malignant glioma heterografts in nu/nu mice.

Since the polyamine metabolism system is very active in proliferative glioma cells, polyamine linked drugs are to be considered as potential antineoplastic agents against malignant gliomas. This study reports the trial of a new compound lineage, the Polyamine Linked Cyclophosphazenes, on human glioblastoma heterografts in nu-nu mice. Two agents are tested: DIAM 3 and DIAM 4. Both show an important antineoplastic action either on a chronic treatment schedule or as single dose. Systemic tolerance is satisfactory.

Further comparison of adjuvants for an inactivated infectious coryza vaccine.

Inactivated infectious coryza vaccines containing different adjuvants were administered to 6-week-old chickens as a single dose containing 10(8) colony-forming units of Haemophilus paragallinarum HP31. After 3 weeks, all chickens were challenged by intrasinus inoculation of HP31. Two vaccines, one containing an aluminum-hydroxide adjuvant and the other a combined aluminum-hydroxide + avridine (a lipoidal amine) adjuvant, were effective. The three remaining vaccines--containing mineral-oil double-emulsion, avridine, or a combination of mineral-oil double-emulsion + avridine--gave much lower levels of protection. No adverse reactions were seen with any vaccine.

The influence of subsequent dehydroepiandrosterone, diaminopropane, phenobarbital, butylated hydroxyanisole and butylated hydroxytoluene treatment on the development of preneoplastic and neoplastic lesions in the rat initiated with di-hydroxy-di-n-propyl nitrosamine.

The comparative modifying potential of dehydroepiandrosterone (DHEA), diaminopropane (DAP), phenobarbital (PB), butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) on the development of lesions initiated by dihydroxy-di-n-propyl nitrosamine (DHPN) in F344 rats were investigated. DHEA, BHA and BHT were all associated with significant reduction in numbers of glutathione-S-transferase P form (GST-P) positive foci in the liver whereas PB brought about their enhanced development. BHT and PB exerted promoting activity on the incidence of thyroid adenomas while DAP similarly increased lung adenoma formation. The results illustrate the advantages to be gained from two stage experiments using broad spectrum carcinogen initiation for comparative analysis of 'modifiers' of the neoplastic process and suggest that studies of enzyme alteration within putative preneoplastic lesions may be directly relevant to elucidation of mechanisms underlying such modification.

Effectiveness of various therapeutic regimens for bovine brucellosis.

Various chemotherapeutic regimens were evaluated in 48 culture-positive dairy cows. Cessation of shedding of Brucella abortus from udder secretions and absence in selected tissues at necropsy were criteria of success. A combination of a long-acting oxytetracycline and streptomycin eliminated Brucella in 10 of 14 (71.4%) cows. Two cows that were retreated with the same regimen also became culture-negative. Other treatment regimens, including the use of liposome-encapsulated antibiotics, were less successful. Serotests were a poor criterion of effectiveness.